Transcription PDF

## Transcription ### What is a Gene? A gene is a specific sequence of nucleotides in the DNA molecule that is concerned by making a special functional RNA. ### Spacer DNA In between genes in DNA, there are sequences of nucleotides that do not code for any RNA. These areas of DNA are called spacer...

## Transcription ### What is a Gene? A gene is a specific sequence of nucleotides in the DNA molecule that is concerned by making a special functional RNA. ### Spacer DNA In between genes in DNA, there are sequences of nucleotides that do not code for any RNA. These areas of DNA are called spacer DNA. ### Transcription For each gene, during transcription, only one strand is transcribed (acting as a template). This strand that is being transcribed is called the **antisense strand**. The resulting RNA is complementary to the antisense strand. The other strand of the DNA is called the **sense strand**. It is not being transcribed and the resulting RNA is identical to this strand. #### Illustration of Transcription A diagram shows the double helix of DNA with the sense and antisense strand labelled. The antisense strand is being transcribed by RNA polymerase to produce the RNA transcript. The direction of transcription is shown with an arrow. ### Major Types of RNA There are four major types of RNA: 1. **r-RNA**: Most abundant (rampant) - 80% 2. **m-RNA**: Most massive 3. **t-RNA**: Smallest (tiny) 4. **s.n. RNA**: Small nuclear RNAs. These are only present in eukaryotes. ### Primary Transcript When RNA is freshly made in the process of transcription, it is called the **primary transcript**. ### Post Transcriptional Modifications The primary transcript undergoes several types of post-transcriptional modifications 1. **Terminal trimming** 2. **Terminal addition** 3. **Internal segment removal** (introns are removed) 4. **Splicing**: Stitching remaining segments together after internal segment removal. 5. **Nitrogenous base modification**: Mostly, this happens to m-RNA. Some of the nitrogenous bases are chemically modified and may become unusual types, other than typical bases for RNA, like adenine, guanine, cytosine and uracil. ### Types of r-RNA There are several types of r-RNA: * **Prokaryotic**: Which has 3 types - 5S, 16S and 23S (S stands for Svedberg Unit). * **Eukaryotic**: Which has 4 types - 5S, 5.8S, 18S, 28S. ### t-RNA Every t-RNA has CCA sequence on 3' end. It is this sequence that attaches and carries an amino acid. ### m-RNA Examples of terminal addition to primary transcript m-RNA: Guanine capping and poly-A tail. * **Guanine capping**: Made of methylated guanine triphosphate 5' to 5' attachment. * **Poly-A tail**: A chain of adenine nucleotides added to the 3' end of the m-RNA. ### Transcription in Prokaryotic genes Prokaryotic RNA polymerase is a very efficient enzyme made of multiple units. The RNAP core enzyme is made of 2 units: * **ω unit**: (omega) unit * **2 β units**: β and β’ The 2 β subunits (β and β’) make the RNAP core enzyme. When the core enzyme binds to the σ factor (sigma factor), only then is the RNA holoenzyme completed and can start to work. * **σ subunit**: (sigma factor) plays a crucial role in recognition of a promoter region of a gene and the holoenzyme's attachment to it. Once the holoenzyme is attached to the promoter, the σ factor dissociates from the core enzyme and moves downstream the DNA unzipping the double helix. * **ω subunit**: (omega) subunit mostly plays a structural role in stabilizing the shape of the core enzyme. * **β and β’ subunits**: Actually perform catalytic function in making phosphodiester bonds between RNA nucleotides and also stabilize RNAP on the DNA strand after σ factor dissociates. **Illustration of the Core Enzyme and Holoenzyme** A simple diagram shows the RNAP which has an α α core enzyme and a σ subunit which is referred to as the holoenzyme. **The Promoter Box** RNAP holoenzyme attaches to and enters a gene in a specific place called a **promoter box**. Usually, the promoter box is on the 5' end (upstream) from the transcribable portion of a gene. Sometimes the promoter box can be on the 3' end (downstream) from it, and very rarely it can be within the transcription area. **Location of the Promoter Box** The location of the promoter box is by convention referred to according to the sense strand, not the template strand. **Diagram of a promoter box** A diagram of a promoter box is shown with the following: * "-35 sequence" is labelled and reads TTGACA. * "Pribnow box" is labelled and reads TATAAT. * Numbers representing the sequence are 5' - 3' and include the -35, Pribnow box, and then up to +3. **RNAP and the Promoter Box** RNAP holoenzyme enters the promoter region either through the Pribnow box or through the -35 sequence. Then the RNAP starts walking downstream but does not start transcribing yet for several nucleotides. The first nucleotide that is transcribed is assigned designation of +1. Nucleotides from +1 back to the promoter box are numbered -1, -2, -3, etc. The promoter box is usually about -10 nucleotides from the +1. **Pribnow box** Around -9 level in the promoter box on the sense strand (not on the template strand) there is a special sequence: **TATAAT**. This zone is called the **Pribnow box**. This is considered as one of the signals for the σ unit to attach to the DNA. Further upstream, at about -35 position, there is another sequence: **TTGACA.** This area is called the **-35 sequence**. RNAP holoenzyme can attach to either of these two sequences in the promoter box. **Proofreading ability** RNA polymerase does **not** have proofreading ability. **Mutations affecting transcription** Mutations in the promoter box area may increase or decrease affinity of RNAP to it and will result in either increase or decrease of that gene expression (rate of transcription).